Abstract Cytokinesis, the physical separation of one cell into two daughter cells, is the final stage of cell division, and although it is the least well understood, it is central to development and tissue homeostasis. Correctly timing cytokinesis so that it occurs only after chromosome replication and segregation is necessary to prevent catastrophic genomic instability, and accordingly, cytokinesis is strictly regulated in concert with other cell cycle events. Using a powerful model organism, the fission yeast Schizosaccharomyces pombe, my lab has conducted pioneering research to identify proteins essential for cytokinesis and to learn how the myriad proteins that comprise the cell division machinery are coordinated to ensure the exquisite spatial and temporal control of cell division. We propose to continue our work pursuing fundamental questions in this field using a multi-disciplinary approach in two directions. In one direction, we will tackle how cytokinesis is entrained with other events of mitosis by investigating the defect that leads to inhibition of cytokinesis when the mitotic spindle is disrupted, how the CK1 enzymes that regulate this branch of the mitotic checkpoint are activated by spindle stress, and how CK1 signalling is integrated with other pathways at spindle poles. Understanding CK1 regulation in the context of the mitotic checkpoint will also establish general mechanisms of regulation for this enzyme family, which are conserved, multifunctional kinases with roles in numerous human diseases. In a second direction, we will advance our understanding of the assembly and architecture of the contractile ring using sophisticated microscopy approaches. We will continue to build our knowledge of the major scaffold of the contractile ring, the F-BAR protein Cdc15, by defining how it oligomerizes on the plasma membrane, and how other contractile ring components are organized on the Cdc15 scaffold. We will also test our hypothesis that multiple cell cycle and polarity kinases inhibit the establishment of the Cdc15 scaffold at inappropriate locations and times, ensuring it only assembles in the cell middle during mitosis. These focused mechanistic studies will be complemented with proteomic and large-scale genetic screens designed to establish a functional interaction network of contractile ring components. Together, these studies will have a major impact for understanding how cytokinesis is orchestrated in eukaryotic species from yeast to humans.